Breast cancer is the most common type of cancer and the second most common cause of cancer-related mortality in women. Cancer cells, including breast cancer, display altered metabolism in order to support their rapid growth and biosynthetic demands. Glycolytic flux is increased, even in the presence of oxygen, in order to conserve necessary building blocks used in synthetic processes. Nutrient sensitive O-GlcNAcylation is the reversible addition of an O-linked GlcNAc moiety to nuclear and cytosolic proteins by the enzyme O-GlcNAc transferase (OGT) [3]. Our lab has previously shown that both O-GlcNAcylation and OGT levels are increased in breast cancer cells compared to normal mammary epithelial cells [12]. Reduction of OGT and O- GlcNAcylation in cancer cells resulted in decreased cancer phenotypes including glycolytic flux and lactate production and ultimately resulted in ER stress induced cell death [13,16]. In this proposal we will elucidate a novel connection between OGT/O-GlcNAc and lipid metabolism in cancer. Preliminary data shows that stable OGT suppression in breast cancer cells results in decreased levels of many lipid metabolites and decreased protein levels of the active form of a key transcriptional regulator of lipid synthesis, Sterol Regulatory Element Binding Protein 1(SREBP-1). Aim #1 will determine whether OGT/O-GlcNAc regulates lipogenesis via SREBP- 1 regulation in breast cancer cells. Aim #2 will determine whether OGT regulation of lipid synthesis is critical for cancer cell survival. The goal of the proposed study is to understand mechanistically how OGT/O-GlcNAc are regulating lipid metabolism in cancer and if this regulation can be exploited for therapeutic purposes in the treatment of breast cancer.